The invention relates to a system for manipulating microparticles in streaming fluids, in particular a procedure for moving microparticles, e.g., biological cells, between various fluids, e.g., for sorting, treatment or confinement purposes, and a microsystem device for implementing the procedure.
Many biological, medical, pharmacological, and even non-biological applications place importance on precise loading with substances and non-contact confinement of microscopic particles, such as biological cells or cell clusters, latex particles or other microbeads in a free liquid. The most frequent method is to grow cells on a solid substrate, which is then rinsed with the required precision using a solution, or to have confinement take place on a sieve or capillary openings. The disadvantage to this procedure is the mechanical surface contact and the difficulty involved in sequentially processing numerous objects in identical fashion. Particular difficulties are encountered in exposing microscopic objects to another solution without surface contact for very short and adjustable times, and then returning them to the original medium. This has previously been achieved at the expense of complicated washing and centrifugation stages.
Use is also made of so-called xe2x80x9claser tweezersxe2x80x9d, with which particles can be held in position in a free solution with micrometer precision, or shifted to a defined extent (see A. Ashkin et al. in xe2x80x9cOptics Lett.xe2x80x9d, Vol. 11, p. 288 (1986)). The disadvantage is that this principle requires a considerable outlay of external equipment, which runs counter to the advantages of system miniaturization and is cost-intensive. In addition, the object is strained in the focal area.
One alternative involves electrical microfield cages, in which microparticles and cells can be held in place similarly to xe2x80x9claser tweezersxe2x80x9d via polarization forces (G. Fuhr et al. xe2x80x9cNaturwiss.xe2x80x9d, Vol. 81, p. 528 (1994)). However, only one solution is situated in such systems, so that the microparticles can only be transferred to another medium via liquid exchange, which requires longer times until the next use and possibly separate cleaning stages. A particle can be held in a confined or parked position by means of a laser tweeter, but it makes no sense from a technical standpoint for several particles. In addition, the object is exposed to a permanent beam load for the time parked.
Magnetically charged particles are transferred from one solution into another in micro systems via magnetic fields or ultrasound sources acting at a right angle to the channels (see G. Blankenstein in xe2x80x9cScientific and Clinical Applications of Magnetic Carriersxe2x80x9d, published by Hafeli et al., Plenum Press New York 1997 (Ch. 16, pp. 233). Both techniques are only suitable for miniaturization under very limited conditions, do not permit focusing the forces acting on the particles, and are difficult to convert in an integrated form using techniques in semiconductor structuring procedures. In addition, this technique is associated with a load with magnetic particles that might be physiologically disruptive for biological objects.
DE-OS 41 43 573 discloses a device for separating mixtures of microscopic particles in a liquid, in which the particles are exposed to electrical fields of traveling waves, during which influence the particles are tapped from a stream of liquid. This device has the following disadvantages. Numerous microelectrodes are required to generate the traveling waves, thus resulting in a complex structure with the respective separate drive circuits. The microelectrodes are situated in an area substantially larger than the particles to be tapped. The traveling waves trigger temperature gradients in the liquid that give rise to disruptive transverse flows. These transverse flows and any other existing flow inhomogeneities prevent the particles from moving along defined paths. To compensate for this locally undefined tapping, it must extend over a relatively wide area in the direction of flow. This in turn results in entire particle groups getting tapped, or the particles must move through the micro system with great distances, thus delaying the processing of large number of particles.
Therefore, the known techniques could previously only be used on a restricted basis, if at all, to transfer microparticles from one liquid to one or more others and back, or to effect a non-contact intermediate storage in a micro system.
Accordingly, the object of the invention is to provide an improved procedure for manipulating microparticles in fluid streams that has an expanded range of application, and in particular can be used serially and parallel at a high velocity, and also enables electrically controllable procedures for the non-contact confinement and transfer of microparticles in various media. The object of the invention is also to provide a device for executing the procedure, which has a simplified design along with a simplified and reliable drive circuit, and is set up to form defined paths of movement for the microparticles to be manipulated.
This object is achieved by a procedure with the features set forth in claim 1 and a device with the features set forth in claim 9. Advantageous embodiments of the invention are described in the dependent claims.
The invention is based on the idea of subjecting microparticles to electromagnetic forces in a streaming fluid. The electromagnetic forces are exerted by at least one electrical field barrier, against which the microparticles are moved with the streaming fluid, and which causes the microparticles to move in a direction deviating from direction of flow. The electrical field barrier is generated, for example, with at least one pair of strip-shaped microelectrodes, which is situated at opposing boundaries of the streaming fluid and exposed to a high-frequency alternating voltage. The selected amplitude for the alternating voltage of field barrier is high enough to prevent the microparticles to be deflected from getting between the electrodes. The fluid with the microparticles suspended therein flows through a channel with at least one lateral hole, to which at least one microparticle is moved along the electric field barrier. The hole is bordered by another channel with a streaming fluid or looped branch (so-called parking loop) of the first channel. The streaming fluids of the respective channels come into contact at the hole. However, the fluids do not become mixed together if the streaming fluid system is implemented with laminar flows. The laminar flows are advantageously implemented in micro systems or with capillary channels. One particular advantage to the invention is that the streaming fluids at the openings between the channels form boundary surfaces that the microparticles to be manipulated can pass through.
The electromagetic field forces are generally exerted via 3-dimensionally distributed electrode arrays through or at the holes between the channels for transferring the objects into one or several adjacent channels or parking loops by applying high-frequency voltages given a permanent hydro-dynamic flow through the system. The electrode arrays acting as deflecting systems can be actuated from a computer, permitting minimal manipulation times in the ms range. The movement can take place in a free solution without any mechanical contact or guidance of the object. The procedure involves no interference and uses the conventional optical measuring methods, thereby avoiding damage to living biological objects, e.g., cells. The time for which the particles reside in the compartments or channel sections can be externally set. Typical path diameters or deflections lie within a range of 50 nm and several 100 xcexcm or more. No feedback check or observation of the objects is required (but can be additionally performed).
Special advantages to the invention are that a highly precise, reliable and rapid particle manipulation is achieved with a relatively simple electrode configuration (in the simplest case: with one pair of electrode strips). Disruptive transverse flows are avoided. The electrodes can be exposed to a sufficiently high alternating voltage, so that the particles reliably stay on the side of the field barrier located upstream, and are routed to the lateral hole. The electrodes have characteristic dimensions smaller than or equal to the dimensions of the particles to be manipulated. Manipulating the particles according to the invention makes it possible to move the particles in and out of the flow, i.e., to also return particles from an adjacent flow.